Inhibition of enterovirus 71 entry by peptides targeting I β-sheet of VP1 protein
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Inhibition of enterovirus 71 entry by peptides targeting I β-sheet of VP1
Ming-Liang He, Ph.DThe Chinese University of Hong Kong
Enterovirus 71 (EV71)
A single stranded, non-enveloped RNA virus (~ 7.5 kb in size);
EV71 infection causes hand-foot-and-mouth disease (HFMD);
Children under 5 are highly susceptible to EV71 and easily develop central nervous system (CNS) symptoms;
As long as it causes CNS infection or symptoms, leading to neurologic sequelae, including neurological disorders, meningitis, and even death.
Hand, Foot & Mouth Disease
an acute viral infection, commonly seen in infants and children
EV71 epidemiology in China
The reported HFMD cases and related death in China from 2007 to 2012.
No drugs, no vaccine available!
Lu J et al., Crit. Rev. Microbiol. 2013
Principle of Antiviral Block viral entry by peptides, small molecules or antibodies;
Block viral replication;
Block the assembly of virions;
Block the virion secretion;
Disturb the cellular signaling, and therefore block one or more processes in the life cycle of virus.
The 3D structure of VP1-VP4 complex of EV71 (PDB ID: 3VBS) virion.
VP1, VP2, VP3 and VP4 were represented in blue, red, green and yellow, respectively. I-β sheet is indicated by a big write arrow.
The VP1 structure
a-helix, D-β and I-β sheet form the canyon, which binds EV71 receptors
The I β sheet is highly conserved among EV71 subtypes and suitable for drug design
Sequence MW PI Antiviral
SP40 (H) QMRRKVELFTYMRFD 2020.4 9.98 Strong
SP45 (β) AEFTFVACTPTGEVV 1935.8 8.68 Weak
SP81 (β) KSKYPLVVRIYMRMK 1912.4 10.56 Very strong
SP81-3 KSKYPLVVRIYMRMKHVRAWI 2675.3 11.12 Very strong
Table 1 The physical and chemical parameters of the synthetic peptides
Peptides target I-β sheet inhibited cytopathic effects caused by EV71 infections
10 uM 50 uM 100 uM 200 uM
Peptides target I-β sheet protected the viability of cells infected by EV71
10 uM 50 uM 100 uM 0
Peptides target I-β sheet inhibited viral reproduction
3CD? 2A, 3C?
IRES5’ 1D 1B 1C 1A 2A 2B 2C 3A 3B 3C 3D
IRES guided translation
VP0 VP3 VP1
2A 2B 2C3A 3C 3D
Proteinase Helicase PolymeraseCleavage
The structure of EV71 genome and its encoding proteins
EV71 pseudovirus system
10 uM 50 uM 100 uM0.00
10 uM 50 uM 100 uM0
Peptides target I-β sheet inhibited viral infectivity
A. Peptides target I-β sheet inhibited the binding of virions to host cells (binding at 4⁰C for 30 min; B. Peptides target I-β sheet inhibited viral entry as indicatedBy reduced translation expression of luciferase reporter.
Sequence Number percentage
KSKYPLVVR I YMRMK 611 61.8%
KSKYPLV I R I YMRMK 359 36.3%
KSKYPLL I R I YMRMK 4 0.4%
KSKYPLV I RMYMRMK 1 0.1%
KSKYPL I VR I YMRMK 2 0.2%
KSKYPL I I R I YMRMK 5 0.5%
KSKYPI VVR I YMRMK 1 0.1%
KSKYPVVVR IYMRMK 1 0.1%
KSKYPLVVRVYMRMK 3 0.3%
KSKYPLVVRTYMRMK 1 0.1%
KSKYPVV I RIYMRMK 1 0.1%
Table 2 The sequences and residue mutation rate of I-β in EV71 circulating worldwide
The sequences of SP81-3 (same as the strain outbreaked in 2008 at Fuyang city, Anhui Province, China; GenBank: ACS12928.1). A total of 989 VP1 sequences was recorded in GeneBank and EMBL database by February 2013. The mutated residues are shown in red color.The approximately accumulated mutation rate: 247L to 247I/V, 0.3%; 248V to 248I, 0.7%; 37.4%; 251I to 251M/V/T, 0.6%.
SP79 RTVGSTKSKYPLVVR -
SP80 GSTKSKYPLVVRIYM +/-
SP81 (β) KSKYPLVVRIYMRMK +++++
SP82 YPLVVRIYMRMKHVR ++++
SP83 VVRIYMRMKHVRAWI ++++
SP84 VVRIYMRMKHVRAWIPRP -
SPC1 AAAVVRIYMRMK ++
SPC2 YPLAAAIYMRMK +
SPC3 YPLVVRAAARMK ++
SPC4 YPLVVRIYMAAA -
SPC5 YPILIRMYMRMK ++++
Table 3 Identification of key residues for EV71 infectivity
SPscr SPC-1 SPC-2 SPC-3 SPC-4 SPC-5 SP810
The antiviral activity of I-β mutants
Arg250, Arg254, Met255 and Lys256 are key residues which locate on theSurface and would involve interaction with EV71 receptors
Conclusion The I β-sheet is an important structure to form the convoy of VP1, which
involves receptor binding;
The residues in I β-sheet are highly conserved among subtypes of EV71, therefore, it is an idea structure for drug target;
Peptide targeting I β-sheet potently inhibited EV71 infections;
Residues Arg250, Arg254, Met255 and Lys256 are critical for virion binding to host cells.